Sepsis-induced acute lung injury is associated with greatly decreased synthesis of surfactant, a stabilizing material comprised mainly of the lipid, phosphatidylcholine (PC). The production of PC is exquisitely controlled by the key surfactant enzyme, CCT. CCT is prone to phosphorylation and proteolytic degradation leading to enzyme inactivation and reduced surfactant availability. Our preliminary data show that in bacterial sepsis models, intracellular calcium triggers kinase-mediated CCT phosphorylation (Aim 1) and F-Box driven ubiquitination and lysosomal degradation of the surfactant enzyme (Aim 2) that impairs surfactant production. Calmodulin, a calcium-regulated sensor appears to protect CCT from these modifications. Thus, we hypothesize that in sepsis, lung injury is mediated, in part, by post-translational events within the CCT primary structure. We will test our hypothesis using state-of-art approaches to identify putative phosphorylation and ubiquitin acceptor sites within the CCT sequence and molecular signatures that govern interactions between F-Box, calmodulin, ubiquitin, and the CCT phosphorylation state. We will also examine the ability of calmodulin to oppose action of F-Box and kinase activity in vitro and in vivo. Our approaches include adenoviral gene transfer of novel surfactant CCT enzymes that are resistant to these post-translational modifications and calmodulin gene transfer in mice after bacterial sepsis. Our long-term goal is to devise suitable pharmaceutical inhibitors of bacterially regulated F-box proteins and kinases or calmodulin activators that might stimulate surfactant production in florid sepsis.